Irreversible binding of tolmetin glucuronic acid esters to albumin in vitro.
Tolmetin glucuronide (TG), extracted and purified from human urine, was incubated with albumin in vitro. The degradation profile and irreversible binding to protein were investigated and kinetic parameters calculated. Standard conditions were as follows: TG, 30 micrograms/ml; human serum albumin (HSA), 3%; pH 7.45; 37 degrees C. Lower pH enhanced TG stability and reduced both the extent and the rate of irreversible binding. HSA also increased TG stability, compared to protein-free buffer, but the opposite was observed with bovine serum albumin (BSA). With BSA, irreversible binding was much less, but the rate of adduct formation was the same as with HSA. Essentially fatty acid free HSA behaved similarly to HSA. Preincubation of HSA with warfarin, or diazepam, or an excess of tolmetin, did not influence irreversible binding significantly. In buffer, acyl migration led predominantly to one isomer. This isomer bound irreversibly to HSA, although more slowly and to a lesser extent than the beta 1-isomer. Incubation of TG with poly-L-lysine also resulted in irreversible binding but to a lesser extent than with HSA. Our results suggest that there is more than one binding mechanism, with the preferential pathway a function of the isomers present and the experimental conditions.
Munafo
A
A
Department of Pharmacy, School of Pharmacy, University of California, San Francisco 94143-0446.
McDonagh
A F
AF
Smith
P C
PC
Benet
L Z
LZ
D000418:Albumins / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007536:Isomerism; D011485:Protein Binding; D014046:Tolmetin / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics / Q000652:urine
D000418:Albumins; C056583:tolmetin glucuronide; D014046:Tolmetin